Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: Illustration of the PiggyBac system for exogenous DNA insertion into the human genome After the co-transfection of the PiggyBac transposase expression plasmid and PiggyBac cloning and expression vector, the expressed PiggyBac transposase cut the Ins, Insulator. ITR, inverted terminal repeat. Adapted from the website of System Biosciences.

Article Snippet: Obtain PiggyBac transposase expression plasmid from System Biosciences (Cat# PB210PA-1).

Techniques: Cotransfection, Expressing, Plasmid Preparation, Cloning

Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: The map of PiggyBac cloning and expression plasmid The schematic illustrates a plasmid backbone containing an ampicillin resistance marker, a multiple cloning site (MCS) for insertion of the desired expression cassette, and an SV40 polyadenylation signal to facilitate proper transcriptional termination.

Article Snippet: Obtain PiggyBac transposase expression plasmid from System Biosciences (Cat# PB210PA-1).

Techniques: Cloning, Expressing, Plasmid Preparation, Marker

Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: FACS analysis showed the GFP+ cell percentage after plasmid transfection The transfection of PiggyBac-SVA-GFP-pA and PiggyBac-SVA-AAA-GFP-pA into HEK293T cells can produce a GFP+ cell population, while the transfection of PiggyBac-SVA-A-GFP-pA and PiggyBac-SVA-AA-GFP-pA cannot, indicating the frame shift of translation.

Article Snippet: Obtain PiggyBac transposase expression plasmid from System Biosciences (Cat# PB210PA-1).

Techniques: Plasmid Preparation, Transfection

Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: Illustration of the PiggyBac system for exogenous DNA insertion into the human genome After the co-transfection of the PiggyBac transposase expression plasmid and PiggyBac cloning and expression vector, the expressed PiggyBac transposase cut the Ins, Insulator. ITR, inverted terminal repeat. Adapted from the website of System Biosciences.

Article Snippet: Plasmid: PiggyBac transposase , System Biosciences , Cat# PB210PA-1.

Techniques: Cotransfection, Expressing, Plasmid Preparation, Cloning

Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: The map of PiggyBac cloning and expression plasmid The schematic illustrates a plasmid backbone containing an ampicillin resistance marker, a multiple cloning site (MCS) for insertion of the desired expression cassette, and an SV40 polyadenylation signal to facilitate proper transcriptional termination.

Article Snippet: Plasmid: PiggyBac transposase , System Biosciences , Cat# PB210PA-1.

Techniques: Cloning, Expressing, Plasmid Preparation, Marker

Journal: STAR Protocols

Article Title: Protocol to identify SINE-VNTR-Alu regulators using genome-wide screening in human K562 cells

doi: 10.1016/j.xpro.2026.104468

Figure Lengend Snippet: FACS analysis showed the GFP+ cell percentage after plasmid transfection The transfection of PiggyBac-SVA-GFP-pA and PiggyBac-SVA-AAA-GFP-pA into HEK293T cells can produce a GFP+ cell population, while the transfection of PiggyBac-SVA-A-GFP-pA and PiggyBac-SVA-AA-GFP-pA cannot, indicating the frame shift of translation.

Article Snippet: Plasmid: PiggyBac transposase , System Biosciences , Cat# PB210PA-1.

Techniques: Plasmid Preparation, Transfection

Journal: bioRxiv

Article Title: Genetic compensation in β-actin mutants occurs independently of mutations that destabilize β-actin mRNA

doi: 10.64898/2026.04.21.719943

Figure Lengend Snippet: (A) Schematic of the doxycycline-inducible construct used for transgene integration and expression. An 8x tetO sequence is positioned upstream of the transgene, IRES, and eGFP, followed by a polyA signal. An EF1a promoter drives rtTA and puromycin resistance expression. The entire cassette is flanked by inverted terminal repeats (grey triangles) for piggyBac transposase-mediated integration. Arrows mark transcription start sites for dox-inducible (8x tetO) and constitutive (EF1a) promoters. Abbreviations: rtTA, reverse tetracycline transactivator; 8x tetO, eight tandem tet operator repeats; IRES, internal ribosome entry site; eGFP, enhanced green fluorescent protein; EF1a, rat elongation factor 1α promoter; pA, polyadenylation sequence; PuroR, puromycin resistance gene; dox, doxycycline. Schematic adapted from Kim et al., 2011. (B) (Top) Schematic of the wild-type ACTB transgene. The sequence corresponds to chr7:5527048–5530601 (hg38). Exons (E1–E6) are represented as rectangles (coding exons are shown as taller boxes than untranslated regions [UTRs]), with introns indicated by connecting lines. The start codon (ATG) is highlighted in green, and the stop codon (TGA) in red. The start and end of the wild-type ACTB mRNA sequence is displayed beneath the gene schematic. (Bottom) Immunoblot for ACTB protein expression in cells before and after 24 hours of doxycycline (dox) treatment. Black arrowhead points to band corresponding to ACTB. ‘M’ indicates the protein molecular weight marker. Ponceau S staining of the membrane is shown as a loading control. (C) RT-qPCR analysis of ACTB , its paralogs ( ACTA1 and ACTG1 ), and a housekeeping gene ( GAPDH ) mRNA expression before (–) or after (+) 24 hours of wild-type ACTB transgene induction by doxycycline. Relative expression was calculated using the 2-ΔΔCt method, normalized to PGK1 mRNA. Error bars denote standard deviation. p-values were calculated with a two-tailed t-test on ΔCt values (gene-of-interest-PGK1). p-value < 0.05 (*), < 0.01 (**), <0.001 (***). (D, E) Same as in (B, C) but for the nonsense mutant ACTB-insPTC transgene, which contains two adjacent PTCs highlighted in red.

Article Snippet: HEK293FT cells were plated at 0.2×10 6 cells/well (12-well plate) and transfected the next day using LipofectamineTM 3000 Transfection Reagent (2 μL, Thermo Fisher Scientific: L3000008) with 500 ng piggyBac transgene plasmid and 500 ng Super PiggyBac transposase plasmid (System Biosciences PB210PA-1).

Techniques: Construct, Expressing, Sequencing, Western Blot, Molecular Weight, Marker, Staining, Membrane, Control, Quantitative RT-PCR, Standard Deviation, Two Tailed Test, Mutagenesis